RESUMO
Accumulating data show that prolylisomerase (Pin1) is overexpressed in human glioblastoma multiforme (GBM) specimens. Therefore, Pin1 inhibitors should be investigated as a new chemotherapeutic drug that may enhance the clinical management of human gliomas. Recently, juglone, a Pin1 inhibitor, was shown to exhibit potent anticancer activity in various tumor cells, but its role in human glioma cells remains unknown. In the present study, we determined if juglone exerts antitumor effects in the U251 human glioma cell line and investigated its potential underlying molecular mechanisms. Cell survival, apoptosis, migration, angiogenesis and molecular targets were identified with multiple detection techniques including the MTT cell proliferation assay, dual acridine orange/ethidium bromide staining, electron microscopy, transwell migration assay, chick chorioallantoic membrane assay, quantitative real-time polymerase chain reaction and immunoblotting. The results showed that 5-20 µM juglone markedly suppressed cell proliferation, induced apoptosis, and enhanced caspase-3 activity in U251 cells in a dose- and time-dependent manner. Moreover, juglone inhibited cell migration and the formation of new blood vessels. At the molecular level, juglone markedly suppressed Pin1 levels in a time-dependent manner. TGF-ß1/Smad signaling, a critical upstream regulator of miR-21, was also suppressed by juglone. Moreover, the transient overexpression of Pin1 reversed its antitumor effects in U251 cells and inhibited juglone-mediated changes to the TGF-ß1/miR-21 signaling pathway. These findings suggest that juglone inhibits cell growth by causing apoptosis, thereby inhibiting the migration of U251 glioma cells and disrupting angiogenesis; and that Pin1 is a critical target for juglone's antitumor activity. The present study provides evidence that juglone has in vitro efficacy against glioma. Therefore, additional studies are warranted to examine the clinical potential of juglone in human gliomas.
Assuntos
Neoplasias Encefálicas/tratamento farmacológico , Glioblastoma/tratamento farmacológico , Naftoquinonas/farmacologia , Animais , Antineoplásicos/farmacologia , Neoplasias Encefálicas/irrigação sanguínea , Neoplasias Encefálicas/enzimologia , Processos de Crescimento Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Embrião de Galinha , Membrana Corioalantoide/efeitos dos fármacos , Glioblastoma/irrigação sanguínea , Glioblastoma/enzimologia , Células Endoteliais da Veia Umbilical Humana , Humanos , Peptidilprolil Isomerase de Interação com NIMA/antagonistas & inibidores , Peptidilprolil Isomerase de Interação com NIMA/metabolismo , Neovascularização Patológica/tratamento farmacológico , Neovascularização Patológica/enzimologia , Transdução de Sinais/efeitos dos fármacosRESUMO
Epidemiological investigations indicate that certain ingredients in tea bricks can antagonize the adverse effects of fluoride. Tea polyphenols (TPs), the most bioactive ingredient in tea bricks, have been demonstrated to be potent bone-supporting agents. ClC7 is known to be crucial for osteoclast (OC) bone resorption. Thus, in this study, we investigated the potential protective effects of TPs against fluorosis using a mouse model and explored the underlying mechanisms with particular focus on ClC7. A total of 40, healthy, 3weekold male C57BL/6 mice were randomly divided into 4 groups (n=10/group) by weight as follows: distilled water (control group), 100 mg/l fluoridated water (F group), water containing 10 g/l TPs (TP group) and water containing 100 mg/l fluoride and 10 g/l TPs (F + TP group). After 15 weeks, and after the mice were sacrificed, the long bones were removed and bone marrow-derived macrophages were cultured ex vivo in order to perform several experiments. OCs were identified and counted by tartrateresistant acid phosphatase (TRAP) staining. The consumption of fluoride resulted in severe fluorosis and in an impaired OC function [impaired bone resorption, and a low mRNA expression of nuclear factor of activated T-cells 1 (NFATc1), ATPase H+ transporting V0 subunit D2 (ATP6v0d2) and osteopetrosisassociated transmembrane protein 1 (Ostm1)]. In the F + TP group, fluorosis was attenuated and OC function was restored, but not the high bone fluoride content. Compared with the F group, mature OCs in the F + TP group expressed higher mRNA levels of ClC7 and Ostm1; the transportation and retaining of Cl was improved, as shown by the fluorescence intensity experiment. On the whole, our findings indicate that TPs mitigate fluorosis in C57BL/6 mice by regulating OC bone resorption. Fluoride inhibits OC resorption by inhibiting ClC7 and Ostm1, whereas TPs attenuate this inhibitory effect of fluoride.
Assuntos
Osso e Ossos/efeitos dos fármacos , Osso e Ossos/metabolismo , Fluorose Dentária/genética , Proteínas de Membrana/genética , Polifenóis/farmacologia , Chá/química , Animais , Biomarcadores , Peso Corporal/efeitos dos fármacos , Reabsorção Óssea/genética , Reabsorção Óssea/metabolismo , Osso e Ossos/patologia , Osso e Ossos/ultraestrutura , Modelos Animais de Doenças , Fluorose Dentária/metabolismo , Fluorose Dentária/patologia , Fluorose Dentária/prevenção & controle , Expressão Gênica , Homeostase , Imuno-Histoquímica , Masculino , Proteínas de Membrana/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Fatores de Transcrição NFATC/genética , Fatores de Transcrição NFATC/metabolismo , Osteoclastos/efeitos dos fármacos , Osteoclastos/metabolismo , Osteoclastos/ultraestrutura , Extratos Vegetais/farmacologia , Substâncias Protetoras/farmacologiaRESUMO
Chronic arsenicosis induced by excessive arsenic intake can cause damages to multi-organ systems, skin cancer and various internal cancers. However, the key metabolic changes and biomarkers which can reflect these changes remain unclear resulting in a lack of effective prevention and treatments. The aim of this study is to determine the impact of chronic arsenic exposure on the metabolism of organism, and find the metabolites changes by using metabolomic techniques. Thirty male Wistar rats were randomly divided into three groups. The arsenite was administered in water, and the doses were 0, 10, and 50mg/L, respectively. The exposure lasted for 6 months. The endogenous metabolite profile of serum was investigated by ultra-performance liquid chromatography coupled with quadrupole time-of-flight tandem mass spectrometry. Partial least squares discriminant analysis (PLS-DA) enabled clusters to be visualized. Nine serum principal metabolites contributing to the clusters were identified, which were CPA (18:2(9Z,12Z)/0:0), LysoPC (14:0), LysoPC (18:4 (6Z,9Z,12Z,15Z)), LysoPC (P-18:0), l-palmitoylcarnitine, LysoPC (20:2(11Z,14Z)) in positive ESI mode and deoxygcholylglycine, LysoPE (0:0/20:2(11Z,14Z)), 15(S)-hydroxyeicosatrienoic acid in negative ESI. These changes of metabolites in rats suggested the changed metabolism in rats exposed to arsenic. These findings may further aid diagnose and serve as targets for therapeutic intervention of arsenicosis.
Assuntos
Intoxicação por Arsênico/metabolismo , Animais , Intoxicação por Arsênico/sangue , Arsenitos/intoxicação , Biomarcadores/sangue , Cromatografia Líquida de Alta Pressão/métodos , Relação Dose-Resposta a Droga , Masculino , Espectrometria de Massas/métodos , Metabolômica , Ratos , Ratos WistarRESUMO
BACKGROUND: Associations between Arg399Gln, Arg194Trp and Arg280His polymorphisms of the XRCC1 gene and risk of differentiated thyroid carcinoma (DTC) have been widely studied but the findings are contradictory. METHODS: We performed a meta-analysis in the present study using STATA 11.0 software to clarify any associations. Electronic literature databases and reference lists of relevant articles revealed a total of 10, 6 and 6 published studies for the Arg399Gln, Arg194Trp and Arg280His polymorphisms, respectively. RESULTS: No significant associations were observed between Arg399Gln and DTC risk in all genetic models within the overall and subgroup meta-analyses, while the Trp/Trp vs Arg/Arg and recessive model of the Arg194Trp polymorphism was associated with DTC susceptibility, and the dominant model of Arg280His polymorphism contributed to DTC susceptibility in Caucasians. CONCLUSIONS: Our meta-analysis suggests that XRCC1 Arg194Trp may be a risk factor for DTC development.
Assuntos
Adenocarcinoma/etiologia , Diferenciação Celular , Proteínas de Ligação a DNA/genética , Polimorfismo Genético/genética , Neoplasias da Glândula Tireoide/etiologia , Predisposição Genética para Doença , Humanos , Prognóstico , Fatores de Risco , Proteína 1 Complementadora Cruzada de Reparo de Raio-XRESUMO
6-Hydroxydopamine (6-OHDA) is a widely used dopaminergic neurotoxin that leads to cell apoptosis in vivo and in vitro, and is a widely accepted experimental model of neurodegeneration in Parkinson's disease. However, the molecular mechanisms responsible for 6-OHDA-induced cell apoptosis are unclear. We found that the treatment of PC12 cells with 6-OHDA resulted in a significant decrease in cell viability and elevated apoptosis as detected by MTT assay, Hoechst 33258 staining, and flow cytometry. In addition, 6-OHDA induced a time-dependent phosphorylation of ERK1/2 at Thr-202/Tyr-204 and of Raf-1 at Ser-338, but a decreased level of Raf-1 phosphorylation at Ser-259. Phosphorylation of ERK1/2 at Thr-202/Tyr-204 and Raf-1 at Ser-338 were inhibited by the Raf-1 inhibitor GW5074, while the ERK1/2 pathway inhibitor U0126 decreased phosphorylation of ERK1/2. Furthermore, 6-OHDA-induced PC12 cells apoptosis was suppressed by GW5074 and U0126. Our results suggest that GW5074 and U0126 act as neuroprotants against 6-OHDA toxicity in PC12 cells by modulating Raf-1/ERK1/2 signaling systems.
Assuntos
Apoptose/efeitos dos fármacos , Butadienos/farmacologia , Inibidores Enzimáticos/farmacologia , Indóis/farmacologia , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Nitrilas/farmacologia , Oxidopamina/antagonistas & inibidores , Fenóis/farmacologia , Proteínas Proto-Oncogênicas c-raf/antagonistas & inibidores , Bisbenzimidazol , Western Blotting , Sobrevivência Celular/efeitos dos fármacos , Citometria de Fluxo , Humanos , Indicadores e Reagentes , Oxidopamina/farmacologia , Células PC12 , Doença de Parkinson/fisiopatologia , FosforilaçãoRESUMO
AIM OF THE STUDY: Although Zuo Gui Yin Decoction has long been used in Traditional Chinese Medicine to treat menopausal symptoms, the underlying mechanism(s) by which these effects are induced remains to be defined. The aim of this study was to investigate the action mechanism of Zuo Gui Yin Decoction on estradiol production in the rat ovary during peri-menopause. MATERIALS AND METHODS: The peri-menopausal animal model was established by natural aging. Peri-menopausal rats were treated by intragastric administration (ig) with low (13.78gkg(-1)), middle (20.67gkg(-1)) or high (31gkg(-1)) dose of Zuo Gui Yin Decoction per day for 8 weeks. At the 8th weekend, the rats were sacrificed for sampling. Estradiol (E(2)) levels in rats' serum were evaluated by radioimmunoassay (RIA). RT-PCR, in situ hybridization and immunohistochemistry were used to determine mRNA and protein expression of relevant genes. RESULTS: Medium- and high-dose of Zuo Gui Yin Decoction could significantly increase serum estradiol concentration, ovarian CYP19 mRNA levels, and P450(arom) protein expression in rats during peri-menopause. Zuo Gui Yin Decoction at three different dosages all could promote FSHR expression and the effect of low-dose was the greatest. Zuo Gui Yin Decoction could elevate LRH-1 and ER(α) expression in a dose dependent manner. CONCLUSIONS: Taken collectively, these findings suggest that Zuo Gui Yin Decoction could promote estradiol production in rat serum during peri-menopausal period through ovarian ER(α)âLRH-1âCYP19 pathway as well as the ovarian classical FSHRâCYP19 mechanism.
Assuntos
Estradiol/biossíntese , Medicina Tradicional Chinesa , Ovário/efeitos dos fármacos , Maturidade Sexual , Animais , Sequência de Bases , Primers do DNA , Feminino , Imuno-Histoquímica , Hibridização In Situ , Ratos , Ratos Wistar , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
OBJECTIVE: The present study aims to investigate the role of protein kinase C delta subtype (PKCdelta) phosphorylation in the process of 6-hydroxydopamine (6-OHDA)-induced dopaminergic cell death, and demonstrate the molecular basis of neurological disorders, such as Parkinson's disease. METHODS: The pheochromocytoma (PC12) cell line was employed in the present study. Cells were treated with 2 mumol/L PKCdelta inhibitor Rottlerin, 10 nmol/L protein kinase C delta subtype (PKCdelta) inhibitor bisindolylmaleimide I, or 5 nmol/L Gö6976 that could specifically inhibit the calcium-dependent PKCdelta isoforms, respectively. PKC activator phorbol-12-myristate-13-acetate (PMA, 100 nmol/L) was also used in this study. All these agents were added to the medium before cells were incubated with 6-OHDA. Cells with no treatment served as control. The cytotoxicity of 6-OHDA was determined by methyl thiazolyl tetrazolium (MTT) reduction assay and PKCdelta phosphorylation levels in various groups were measured by western blotting. RESULTS: Bisindolylmaleimide I and Gö6976 exerted no significant attenuation on the cytotoxicity of 6-OHDA, nor any effects on PKCdelta phosphorylation in PC12 cells. However, Rottlerin could inhibit the phosphorylation of PKCdelta and attenuate 6-OHDA-induced cell death, and the cell viability was raised to (69.6+/-2.63)% of that in control group (P<0.05). In contrast, PMA induced a significant increase in PKCdelta phosphorylation and also strengthened the cytotoxic effects of 6-OHDA. The cell viability of PMA-treated PC12 cells decreased to (49.8+/-5.06)% of that in control group (P<0.001). CONCLUSION: Rottlerin can protect PC12 cells from cytotoxicity of 6-OHDA probably by inhibiting PKCdelta phosphorylation. The results suggest that PKCdelta may be a key regulator of neuron loss in Parkinson's disease.
Assuntos
Acetofenonas/farmacologia , Adrenérgicos/toxicidade , Benzopiranos/farmacologia , Dopamina/metabolismo , Inibidores Enzimáticos/farmacologia , Oxidopamina/toxicidade , Animais , Carbazóis/farmacologia , Sobrevivência Celular/efeitos dos fármacos , Relação Dose-Resposta a Droga , Interações Medicamentosas , Ativação Enzimática/efeitos dos fármacos , Regulação Enzimológica da Expressão Gênica/efeitos dos fármacos , Indóis/farmacologia , Maleimidas/farmacologia , Células PC12 , Ésteres de Forbol/farmacologia , Fosforilação/efeitos dos fármacos , Proteína Quinase C/metabolismo , Ratos , Fatores de TempoRESUMO
OBJECTIVES: An abundance of X chromosomes in testicular germ cell tumors (TGCTs), and a candidate TGCTs susceptibility gene (TGCT1) on Xq27 highlight the potential involvement of X chromosomes in TGCT pathogenesis. However, the TGCT1 on Xq27 has so far not been identified. We hypothesized that a somatic mutation of dbl oncogene on Xq27 may play a role for the development of TGCTs. METHODS: We have screened 41 TGCT tissues for dbl mutations using single-strand conformation polymorphism (SSCP) analysis. These tissues are composed of 25 seminomatous TGCTs tissues and 16 non-seminomatous TGCTs tissues, including two cases with a rhabdomyosarcoma component. RESULTS: Somatic mutations were not detected in the 25 exons of dbl in these TGCTs. However, we found a rare single nucleotide polymorphism (SNP) (T to C nucleotide change) within intron 22 in one out of the 41 TGCTs cases (2%). Furthermore, the sample with the rare SNP was identified as the sole TGCTs case associated with bilateral undescended testis in our series. CONCLUSIONS: Our results indicate that proto-oncogene dbl is not a major target for sporadic TGCTs. However, the rare SNP in dbl may affect the susceptibility to undescended testis. Determining the frequency of this SNP in patients with various types of undescended testis in different ethnic groups is a warranted study.
Assuntos
Criptorquidismo/genética , Fatores de Troca do Nucleotídeo Guanina/genética , Neoplasias Embrionárias de Células Germinativas/genética , Proteínas Proto-Oncogênicas/genética , Neoplasias Testiculares/genética , Cromossomos Humanos X , Análise Mutacional de DNA , Predisposição Genética para Doença , Humanos , Masculino , Polimorfismo de Nucleotídeo Único , Polimorfismo Conformacional de Fita Simples , Proto-Oncogene MasRESUMO
Single cell gel electrophoresis assay (SCGE), also named as alkaline comet assay, was a simple, rapid and sensitive method to evaluate DNA damage. In this study SCGE technique was used to monitor DNA damage difference in tumor patients caused by chemotherapy, DNA damage distribution frequency and DNA damage characters were analyzed by komet image analysis system (KIAS). The results showed that cyclophosphamide greatly caused DNA damage in lymphocytes of tumor patients. There was significant difference of peripheral blood lymphocyte DNA damage between tumor patients and healthy controls. Tail length of lymphocytes were 33.69 +/- 7.56 micro m, and tail DNA% we re 31.51 +/- 5.4 6% in 10 cancer patients treated with cyclophosphamide, while Tail length were 1 6.2 +/- 1.5 micro m and tail DNA% were 7.46 +/- 1.15% in healthy controls. there was great significant difference on tail length and tail DNA% values between cancer patients and healthy controls (P < 0.01). In conclusion, the successful measurement of DNA damage caused by Cyclophosphamide treatment means that the alkaline comet assay as a valuable tool can be very useful in cancer epideminology study, and also be valuable to evaluate DNA damage status of patients in clinic.